Constant pH

What am I possibly hoping to achieve?

Bas Rustenburg // Chodera lab

#pHallthetime

Stardate 93387.66

Disclaimer

For the purpose of this presentation, I've experimented with multiple ways to provide a more interactive experience. Feedback is welcome.

Live web-cast

Follow the slides live on your own computer using this url:

#pHallthetime

http://slides.com/basrustenburg/deck/live

Why am I interested in constant-pH work?

Protonation state effects

Neeb et al. J. Med. Chem., 2014, 57 (13), pp 5554–5565

There is no a priori way to know every relevant protonation state of a ligand or protein, and how relevant they are to the binding affinity.

The standard state approach

Typically, one only simulates a single protonation state.
For proteins at neutral pH, only histidine is closely looked at.

Problems that...

If reality has a mixture of protonation states, we're ignoring their contributions to the free energy of the system
If the most relevant protonation state changes upon binding, we simulate the wrong state

Bonus question

What kinase inhibitor is this?

It's....

Lapatinib!

#ofcourseitslapatinib

The standard approach is ignoring contributions of unknown magnitude, for imatinib and Abl there is clear indication of the relevance

Aleksandrov, A and Simonson, T J Comput Chem 31,7, pp. 1550–1560 (2010)

Kinase inhibitors

pKa predictions using EPIK suggest that many kinase inhibitors have more than one accessible protonation state at pH 7.4

Figure by: John & Julie

What is...

constant-pH?

#pHallthetime

And how does it solve our problems?

Constant PH

Allow for the exchange of protons with an external proton "bath", which maintains a constant pH.

a_{H^+} = \exp \left( \frac{\mu_{H^+} - \mu^\ominus_{H^+} } {RT} \right)

a_{H^+} = \exp \left( \frac{\mu_{H^+} - \mu^\ominus_{H^+} } {RT} \right)

pH = - \log_{10} (a_{H^+}) \approx - \log_{10} [H^+]

pH = - \log_{10} (a_{H^+}) \approx - \log_{10} [H^+]

Constant PH

In implicit solvent

Take a reference state with a known pKa, such as a free dipeptide or capped peptide in solution
ΔGelec difference between the electrostatic potential for the current protonation state and proposed state

\Delta G = k_{\mathrm{B}T} \left(\mathrm{pH - pK_{a,ref}} \right) \ln 10 + \Delta G_{\mathrm{elec}} - \Delta G_{\mathrm{elec,ref}}

\Delta G = k_{\mathrm{B}T} \left(\mathrm{pH - pK_{a,ref}} \right) \ln 10 + \Delta G_{\mathrm{elec}} - \Delta G_{\mathrm{elec,ref}}

Mongan, Case, and McCammon • Journal of Computational Chemistry

Volume 25, Issue 16, pages 2038–2048, December 2004

Constant PH

In implicit solvent

(ΔGelec) take difference between the electrostatic potential for the current protonation state and proposed state
Accept using

\Delta G = k_{\mathrm{B}T} \left(\mathrm{pH - pK_{a,ref}} \right) \ln 10 + \Delta G_{\mathrm{elec}} - \Delta G_{\mathrm{elec,ref}}

\Delta G = k_{\mathrm{B}T} \left(\mathrm{pH - pK_{a,ref}} \right) \ln 10 + \Delta G_{\mathrm{elec}} - \Delta G_{\mathrm{elec,ref}}

Mongan, Case, and McCammon • Journal of Computational Chemistry

Volume 25, Issue 16, pages 2038–2048, December 2004

P = \begin{cases} 1 & \quad \text{if } \Delta G \leq 0 \\ e^{-\Delta G / k_{BT}} & \quad \text{if } \Delta G > 0 \end{cases}

P = \begin{cases} 1 & \quad \text{if } \Delta G \leq 0 \\ e^{-\Delta G / k_{BT}} & \quad \text{if } \Delta G > 0 \end{cases}

Openmm-ConstpH

Currently works for implicit solvent only
Systems are prepared using Ambertools, including tleap, cpinutils
Residue names need to be predefined (tough for ligands)
Have you ever seen a cpinfile?

https://github.com/choderalab/openmm-constph

#pHallthetime

Openmm-ConstpH

Example of a cpin file

&CNSTPH
 CHRGDAT=-0.4157,0.2719,0.0145,0.0779,-0.0398,-0.0173,-0.0173,0.0136,-0.0425,
 -0.0425,0.8054,-0.8188,-0.8188,0.0,0.5973,-0.5679,0.0,0.0,0.0,-0.4157,0.2719,
 0.0145,0.0779,-0.0071,0.0256,0.0256,-0.0174,0.043,0.043,0.6801,-0.5838,-0.6511,
 0.4641,0.5973,-0.5679,0.0,0.0,0.0,-0.4157,0.2719,0.0145,0.0779,-0.0071,0.0256,
 0.0256,-0.0174,0.043,0.043,0.6801,-0.5838,-0.6511,0.0,0.5973,-0.5679,0.4641,
 0.0,0.0,-0.4157,0.2719,0.0145,0.0779,-0.0071,0.0256,0.0256,-0.0174,0.043,0.043,
 0.6801,-0.6511,-0.5838,0.0,0.5973,-0.5679,0.0,0.4641,0.0,-0.4157,0.2719,0.0145,
 0.0779,-0.0071,0.0256,0.0256,-0.0174,0.043,0.043,0.6801,-0.6511,-0.5838,0.0,
 0.5973,-0.5679,0.0,0.0,0.4641,-0.3479,0.2747,-0.24,0.1426,-0.0094,0.0362,
 0.0362,0.0187,0.0103,0.0103,-0.0479,0.0621,0.0621,-0.0143,0.1135,0.1135,
 -0.3854,0.34,0.34,0.34,0.7341,-0.5894,-0.3479,0.2747,-0.24,0.1426,-0.10961,
 0.034,0.034,0.06612,0.01041,0.01041,-0.03768,0.01155,0.01155,0.32604,-0.03358,
 -0.03358,-1.03581,0.0,0.38604,0.38604,0.7341,-0.5894,-0.4157,0.2719,0.0341,
 0.0864,-0.1783,-0.0122,-0.0122,0.7994,-0.8014,-0.8014,0.0,0.5973,-0.5679,0.0,
 0.0,0.0,-0.4157,0.2719,0.0341,0.0864,-0.0316,0.0488,0.0488,0.6462,-0.5554,
 -0.6376,0.4747,0.5973,-0.5679,0.0,0.0,0.0,-0.4157,0.2719,0.0341,0.0864,-0.0316,
 0.0488,0.0488,0.6462,-0.5554,-0.6376,0.0,0.5973,-0.5679,0.4747,0.0,0.0,-0.4157,
 0.2719,0.0341,0.0864,-0.0316,0.0488,0.0488,0.6462,-0.6376,-0.5554,0.0,0.5973,
 -0.5679,0.0,0.4747,0.0,-0.4157,0.2719,0.0341,0.0864,-0.0316,0.0488,0.0488,
 0.6462,-0.6376,-0.5554,0.0,0.5973,-0.5679,0.0,0.0,0.4747,-0.4157,0.2719,
 -0.0014,0.0876,-0.0152,0.0295,0.0295,-0.0011,-0.1906,0.1699,-0.2341,0.1656,
 0.3226,-0.5579,0.3992,-0.2341,0.1656,-0.1906,0.1699,0.5973,-0.5679,-0.4157,
 0.2719,-0.0014,0.0876,-0.0858,0.019,0.019,-0.213,-0.103,0.132,-0.498,0.132,
 0.777,-0.814,0.0,-0.498,0.132,-0.103,0.132,0.5973,-0.5679,
 PROTCNT=0,1,1,1,1,3,2,0,1,1,1,1,1,0,
 RESNAME='System: Unknown','Residue: GL4 7','Residue: LYS 13','Residue: AS4 18',
 'Residue: TYR 20','Residue: TYR 23','Residue: LYS 33','Residue: GL4 35',
 'Residue: AS4 48','Residue: AS4 52','Residue: TYR 53','Residue: AS4 66',
 'Residue: AS4 87','Residue: LYS 96','Residue: LYS 97','Residue: AS4 101',
 'Residue: LYS 116','Residue: AS4 119',
 RESSTATE=0,0,0,0,0,0,0,0,0,0,0,0,0,0,0,0,0,
 STATEINF(0)%FIRST_ATOM=102, STATEINF(0)%FIRST_CHARGE=0, 
 STATEINF(0)%FIRST_STATE=0, STATEINF(0)%NUM_ATOMS=19, STATEINF(0)%NUM_STATES=5, 
 STATEINF(1)%FIRST_ATOM=187, STATEINF(1)%FIRST_CHARGE=95, 
 STATEINF(1)%FIRST_STATE=5, STATEINF(1)%NUM_ATOMS=22, STATEINF(1)%NUM_STATES=2, 
 STATEINF(2)%FIRST_ATOM=276, STATEINF(2)%FIRST_CHARGE=139, 
 STATEINF(2)%FIRST_STATE=7, STATEINF(2)%NUM_ATOMS=16, STATEINF(2)%NUM_STATES=5, 
 STATEINF(3)%FIRST_ATOM=306, STATEINF(3)%FIRST_CHARGE=219, 
 STATEINF(3)%FIRST_STATE=12, STATEINF(3)%NUM_ATOMS=21, 
 STATEINF(3)%NUM_STATES=2, STATEINF(4)%FIRST_ATOM=358, 
 STATEINF(4)%FIRST_CHARGE=219, STATEINF(4)%FIRST_STATE=12, 
 STATEINF(4)%NUM_ATOMS=21, STATEINF(4)%NUM_STATES=2, 
 STATEINF(5)%FIRST_ATOM=500, STATEINF(5)%FIRST_CHARGE=95, 
 STATEINF(5)%FIRST_STATE=5, STATEINF(5)%NUM_ATOMS=22, STATEINF(5)%NUM_STATES=2, 
 STATEINF(6)%FIRST_ATOM=542, STATEINF(6)%FIRST_CHARGE=0, 
 STATEINF(6)%FIRST_STATE=0, STATEINF(6)%NUM_ATOMS=19, STATEINF(6)%NUM_STATES=5, 
 STATEINF(7)%FIRST_ATOM=741, STATEINF(7)%FIRST_CHARGE=139, 
 STATEINF(7)%FIRST_STATE=7, STATEINF(7)%NUM_ATOMS=16, STATEINF(7)%NUM_STATES=5, 
 STATEINF(8)%FIRST_ATOM=789, STATEINF(8)%FIRST_CHARGE=139, 
 STATEINF(8)%FIRST_STATE=7, STATEINF(8)%NUM_ATOMS=16, STATEINF(8)%NUM_STATES=5, 
 STATEINF(9)%FIRST_ATOM=805, STATEINF(9)%FIRST_CHARGE=219, 
 STATEINF(9)%FIRST_STATE=12, STATEINF(9)%NUM_ATOMS=21, 
 STATEINF(9)%NUM_STATES=2, STATEINF(10)%FIRST_ATOM=1028, 
 STATEINF(10)%FIRST_CHARGE=139, STATEINF(10)%FIRST_STATE=7, 
 STATEINF(10)%NUM_ATOMS=16, STATEINF(10)%NUM_STATES=5, 
 STATEINF(11)%FIRST_ATOM=1326, STATEINF(11)%FIRST_CHARGE=139, 
 STATEINF(11)%FIRST_STATE=7, STATEINF(11)%NUM_ATOMS=16, 
 STATEINF(11)%NUM_STATES=5, STATEINF(12)%FIRST_ATOM=1446, 
 STATEINF(12)%FIRST_CHARGE=95, STATEINF(12)%FIRST_STATE=5, 
 STATEINF(12)%NUM_ATOMS=22, STATEINF(12)%NUM_STATES=2, 
 STATEINF(13)%FIRST_ATOM=1468, STATEINF(13)%FIRST_CHARGE=95, 
 STATEINF(13)%FIRST_STATE=5, STATEINF(13)%NUM_ATOMS=22, 
 STATEINF(13)%NUM_STATES=2, STATEINF(14)%FIRST_ATOM=1536, 
 STATEINF(14)%FIRST_CHARGE=139, STATEINF(14)%FIRST_STATE=7, 
 STATEINF(14)%NUM_ATOMS=16, STATEINF(14)%NUM_STATES=5, 
 STATEINF(15)%FIRST_ATOM=1767, STATEINF(15)%FIRST_CHARGE=95, 
 STATEINF(15)%FIRST_STATE=5, STATEINF(15)%NUM_ATOMS=22, 
 STATEINF(15)%NUM_STATES=2, STATEINF(16)%FIRST_ATOM=1810, 
 STATEINF(16)%FIRST_CHARGE=139, STATEINF(16)%FIRST_STATE=7, 
 STATEINF(16)%NUM_ATOMS=16, STATEINF(16)%NUM_STATES=5, 
 STATENE=0,14.4542090223,14.4542090223,14.4542090223,14.4542090223,
 -0.946105574619,0,0,32.3880313021,32.3880313021,32.3880313021,32.3880313021,0,
 -78.310003685,
 TRESCNT=17,
/

https://github.com/choderalab/openmm-constph

Openmm-ConstpH

Amber files are read into openMM using a MonteCarloTitration driver object

    # Parameters.
    niterations = 500 # number of dynamics/titration cycles to run
    nsteps = 500 # number of timesteps of dynamics per iteration
    temperature = 300.0 * units.kelvin
    timestep = 1.0 * units.femtoseconds
    collision_rate = 9.1 / units.picoseconds
    pH = 7.0

    # Filenames.
    prmtop_filename = 'amber-example/prmtop'
    inpcrd_filename = 'amber-example/min.x'
    cpin_filename = 'amber-example/cpin'
   
    # Load the AMBER system.
    import simtk.openmm.app as app
    inpcrd = app.AmberInpcrdFile(inpcrd_filename)
    prmtop = app.AmberPrmtopFile(prmtop_filename)
    system = prmtop.createSystem(implicitSolvent=app.OBC2, nonbondedMethod=app.NoCutoff, constraints=app.HBonds)

    # Instantiate system driver
    mc_titration = MonteCarloTitration(system, temperature, pH, prmtop, cpin_filename, debug=True)

    # Create integrator and context.
    platform_name = 'OpenCL'
    platform = openmm.Platform.getPlatformByName(platform_name)
    integrator = openmm.LangevinIntegrator(temperature, collision_rate, timestep)
    context = openmm.Context(system, integrator, platform)
    context.setPositions(inpcrd.getPositions())


    # Etc. etc.

https://github.com/choderalab/openmm-constph

MonteCarloTitration

Current structure of the MonteCarloTitration driver contains methods such as

def addTitratableGroup(self, atom_indices):
   """
   Define a new titratable group.
   
   ARGUMENTS
   atom_indices (list of int) - the atom indices defining the titration group
   """

def addTitrationState(self, titration_group_index, pKref, relative_energy, charges, proton_count):
    """
    Add a titration state to a titratable group.
    """

def update(self, context):
    """
    Perform a Monte Carlo update of the titration state.
    """

Future plans

How to improve OpenMM constantpH

#pHallthetime

Future plans: ConstpH

Get rid of the CPIN file in favor of forcefield XML flavored format

<TitratableResidues>

<TitratableResidue name="ASP">
<TitrationState index=0>
    <NonbondedForce coulomb14scale="0.833333" lj14scale="0.5">
    <Atom type="ASP-OD1" charge="-0.45" sigma="xxxxx" epsilon="xxxxxx"/>
    <Atom type="ASP-OD2" charge="-0.288" sigma="xxxxx" epsilon="xxxxxx"/>
    <Atom type="ASP-HD2" charge="0.408" sigma="xxxxx" epsilon="xxxxxx"/>
    <Atom type="ASP-CG" charge="0.33" sigma="xxxxx" epsilon="xxxxxx"/>    
    </NonbondedForce>
    
  </TitrationState>
  <TitrationState index=1>
    <NonbondedForce coulomb14scale="0.833333" lj14scale="0.5">
    <Atom type="ASP-OD1" charge="-0.635" sigma="xxxxx" epsilon="xxxxxx"/>
    <Atom type="ASP-OD2" charge="-0.635" sigma="xxxxx" epsilon="xxxxxx"/>
    <!-- Prevent divide by zero! -->
    <Atom type="ASP-HD2" charge="-0.0" sigma="0.000" epsilon="0.000"/> 
    <Atom type="ASP-CG" charge="0.27" sigma="xxxxx" epsilon="xxxxxx"/>    
    </NonbondedForce>
  </TitrationState>
</TitratableResidue>

</TitratableResidues>

#pHallthetime

Expand to incorporate LJ parameters
- Currently, just charges are supported
Better support for ligands
Explicit solvent using NCMC & counter ions
- Inserting ions is unlikely to be accepted because of close contacts, water rearrangement, and more...
Coordinated changes (pairs of residues, etc.) to increase acceptance probability
- Pick pairs at random
- Find a way to pick pairs close together that satisfies detailed balance

Future plans: ConstpH

Benchmark pKa calculation tools to find the most reliable reference values for ligands
Investigate the need for charge corrections in PME calculations (since we are modifying system charges)
Investigate the need to go into the OpenMM C++/Cuda layer for more efficient calculations

Future plans: ConstpH

#pHallthetime

Systems of interest

Kinases & FDA approved inhibitors
Czodrowski et al. protease inhibitors

#pHallthetime

Systems of interest

Kinases & FDA approved inhibitors
Czodrowski et al. protease inhibitors
Lactate dehydrogenase A (LDHA)

#pHallthetime

Kinase inhibitors

pKa predictions using EPIK suggest that many kinase inhibitors have more than one accessible protonation state at pH 7.4

Figure by: John & Julie

Lactate Dehydrogenase A

PDB: 1I10

Discovered a pH dependent effect to the reaction rate
Continued collaboration with Andrew Intlekofer (Thompson lab)

Intlekofer et al. Cell Metabolism 22, 304–311, August 4, 2015

Lactate Dehydrogenase A

PDB: 1I10

Lactate Dehydrogenase A

PDB: 1I10

Thanks for your attention

Confucius says:

"Transfer some protons today!"

pH, all the time

By Bas Rustenburg

pH, all the time

Constant pH presentation // Chodera lab

Bas Rustenburg

ze_europs

Constant pH

Disclaimer

Live web-cast

Why am I interested in constant-pH work?

Protonation state effects

The standard state approach

Problems that...

Bonus question

It's....

More problems with the Standard aproach

Kinase inhibitors

What is...

constant-pH?

Constant PH

Constant PH

Constant PH

Openmm-ConstpH

Openmm-ConstpH

Openmm-ConstpH

MonteCarloTitration

Future plans

How to improve OpenMM constantpH

Future plans: ConstpH

Future plans: ConstpH

Future plans: ConstpH

Systems of interest

Systems of interest

Kinase inhibitors

Lactate Dehydrogenase A

Lactate Dehydrogenase A

Lactate Dehydrogenase A

Thanks for your attention

Confucius says:

"Transfer some protons today!"

pH, all the time

More from Bas Rustenburg